The PI discovered the first metastasis suppressor gene, nm23. Basic and translational research has investigated the role of Nm23 in the regulation of tumor metastasis. Eleven transfection studies have documented that overexpression of Nm23 in various tumor cell lines resulted in a 50-90% decrease in tumor metastatic potential in vivo. The biochemical mechanism whereby Nm23 suppresses metastatic potential is under investigation, based on microarray analysis of control- and Nm23-H1 transfectants. We have identified several genes that were down-regulated in nm23 transfectants, but not in transfectants of mutant forms of nm23 which failed to suppress in vitro motility, including c-met, CTGF, EDG2, FZD1, L1CAM, NETO2, PTN and SMOH. Confirmatory experiments included the down-regulation of the gene set in: (a) wild type as opposed to nm23-M1 knockout mouse hepatocellular carcinoma tissues; (b)independent MDA-MB-435 breast carcinoma transfectants; (c) nm23-H1 transfectants of another breast carcinoma cell line, MDA-MB-231; (d) an cohort of breast carcinomas, stratified by nm23 expression level. Transfection of each of the genes into nm23-suppressed MDA-MB-435 cells revealed that only EDG2,and to a lesser extent c-met, overcame nm23-H1 inhibition of motility. The data suggest that Nm23 inhibits tumor motility by down regulating the receptors to widely available factors such as LPA (EDG2) and HGF (Met). Metastasis experiments using EDG2 transfectants are underway. Translational research on Nm23 proposed that elevation of Nm23 expression in micrometastatic or overtly metastatic breast or other carcinomas may limit colonization, motility and de-differentiation, with a clinical benefit. Analysis of the nm23-H1 promoter revealed a 400 bp region which controlled expression, and contained a cassette of transcription factors regulated by a glucocorticoid response element (GRE). Deletion studies showed that these sites were functional in regulating nm23-H1 transcription. Medroxyprogesterone acetate (MPA), an unusual agonist for GR, as well as the androgen receptor and progesterone receptor, elevated Nm23-H1 expression of breast carcinoma cell lines in vitro. MPA acted via a post-transcriptional mechanism using the GR, at pharmacologic doses. We have reported preclinical experiments to determine if MPA can halt metastatic colonization. Mice were injected iv with metastatic human MDA-MB-231 breast carcinoma cells, and permitted to develop micrometastases for one month. Mice were then randomized to vehicle or MPA, the latter given in a one month induction and subsequent bimonthly maintenance dose. Mice receiving MPA had significantly fewer gross metastases in the lung, a smaller proportion of mice with metastases and smaller metastases. Immunohistochemistry revealed that MPA treated mice had a greater proportion of pulmonary metastases with high Nm23 expression. Side effects included weight gain, but no effects on bone mineral density or mammary histology. The data indicate that agents elevating metastsis suppressor gene expression may be effective against metastatic colonization. A Phase I trial of MPA will open at Indiana University, (PI Kathy Miller) in 2006, funded by an Avon-NCI grant to Dr. Miller. Our laboratory and that of Dr. Merino will participate in the trial.